A candidate ion-retaining state in the inward-facing conformation of sodium/galactose symporter: Clues from atomistic simulations

The recent Vibrio parahaemolyticus sodium/galactose (vSGLT) symporter crystal structure captures the protein in an inward-facing substrate-bound conformation, with the sodium ion placed, by structural alignment, in a site equivalent to the Na2 site of the leucine transporter (LeuT). A recent study, based on molecular dynamics simulations, showed that the sodium ion spontaneously leaves its initial position diffusing outside vSGLT, toward the intracellular space. This suggested that the crystal structure corresponds to an ion-releasing state of the transporter. Here, using metadynamics, we identified a more stable Na+ binding site corresponding to a putative ion-retaining state of the transporter. In addition, our simulations, consistently with mutagenesis studies, highlight the importance of D189 that, without being one of the NA(+)-coordinating residues, regulates its binding/release

Locally accessible conformations of proteins: multiple molecular dynamics simulations of crambin.

Multiple molecular dynamics (MD) simulations of crambin with different initial atomic velocities are used to sample conformations in the vicinity of the native structure. Individual trajectories of length up to 5 ns sample only a fraction of the conformational distribution generated by ten independent 120 ps trajectories at 300 K. The backbone atom conformational space distribution is analyzed using principal components analysis (PCA). Four different major conformational regions are found. In general, a trajectory samples only one region and few transitions between the regions are observed. Consequently, the averages of structural and dynamic properties over the ten trajectories differ significantly from those obtained from individual trajectories. The nature of the conformational sampling has important consequences for the utilization of MD simulations for a wide range of problems, such as comparisons with X-ray or NMR data. The overall average structure is significantly closer to the X-ray structure than any of the individual trajectory average structures. The high frequency (less than 10 ps) atomic fluctuations from the ten trajectories tend to be similar, but the lower frequency (100 ps) motions are different. To improve conformational sampling in molecular dynamics simulations of proteins, as in nucleic acids, multiple trajectories with different initial conditions should be used rather than a single long trajectory

Influence of the heme pocket conformation on the structure and vibrations of the Fe-CO bond in myoglobin: a QM/MM density functional study.

The influence of the distal pocket conformation on the structure and vibrations of the heme-CO bond in carbonmonoxy myoglobin (MbCO) is investigated by means of hybrid QM/MM calculations based on density functional theory combined with a classical force field. It is shown that the heme-CO structure (QM treated) is quite rigid and not influenced by the distal pocket conformation (MM treated). This excludes any relation between FeCO distortions and the different CO absorptions observed in the infrared spectra of MbCO (A states). In contrast, both the CO stretch frequency and the strength of the CO...His64 interaction are very dependent on the orientation and tautomerization state of His64. Our calculations indicate that the CO...N(epsilon) type of approach does not contribute to the A states, whereas the CO...H-N(epsilon) interaction is the origin of the A(1) and A(3) states, the His64 residue being protonated at N(epsilon). The strength of the CO...His64 interaction is quantified, in comparison with the analogous O(2)...His64 interaction and with the observed changes in the CO stretch frequency. Additional aspects of the CO...His64 interaction and its biological implications are discussed

Unexpected Roles of Guest Polarizability and Maximum Hardness, and of Host Solvation in Supramolecular Inclusion Complexes: A Dual Theoretical and Experimental Study

The origin of differential binding affinity and structural recognition between the inclusion complexes of cyclobis(paraquat-p-phenylene), 1 4+, and 1,4-substituted phenyl or 4,4‘-substituted biphenyl derivatives has been jointly determined by spectrometric techniques and ab initio and semiempirical molecular orbital methods. The unusual boxed geometry and tetracationic charge distribution in 1 4+ are key molecular features which produce strong intermolecular interactions with guest and solvent molecules. Solvation was addressed by including up to 12 acetonitrile molecules in the theoretical model, which realigned the predicted gas-phase supramolecular structures and energies into excellent agreement with experiment. The computed complexation enthalpies, ΔH bind, from the semiempirical molecular orbital PM3 method are on average within 1 kcal/mol of the experimental free energy binding data collected from absorption spectroscopy in acetonitrile. In addition, the computed geometric penetration and positioning of 1 4+/benzidine and 1 4+/4,4‘-biphenol complexes are consistent with that reported from NMR NOE data. The partitioning of self-consistent field complexation energies from both classical and quantum forces has been determined by using Morokuma's variational energy decomposition technique. It was determined that the primary basis for the molecular recognition between 1,4-substituted phenyl guests and 1 4+ is short-range stabilizing electrostatic forces complemented by small amounts of polarizability and charge-transfer. In contrast, the recognition force between 4,4‘-substituted biphenyl guests and 1 4+ is dominated by polarizability with a small contribution from electrostatics. Therefore, the balance between molecular polarizability and electrostatics controls the differential binding affinity and structural recognition with 1 4+. For the first time, we report that individual molecular properties of substituted guests correlate with the binding energies of corresponding 1 4+ inclusion complexes. Direct correlations between the 1 4+ binding energies and the computed molecular polarizability, maximum hardness, softness, and electronegativity of the guest have been identified. It is now plausible to consider the design and construction of new supramolecular assemblies based upon a few select molecular properties of the constituent molecules

Delta SARS-CoV‑2 s2m Structure, Dynamics, and Entropy: Consequences of the G15U Mutation

Bioinformatic analysis of the Delta SARS-CoV-2 genome reveals a single nucleotide mutation (G15U) in the stem-loop II motif (s2m) relative to ancestral SARS-CoV-2. Despite sequence similarity, unexpected differences between SARS-CoV-2 and Delta SARS-CoV-2 s2m homodimerization experiments require the discovery of unknown structural and thermodynamic changes necessary to rationalize the data. Using our reported SARS-CoV-2 s2m model, we induced the G15U substitution and performed 3.5 microseconds of unbiased molecular dynamics simulation at 283 and 310 K. The resultant Delta s2m adopted a secondary structure consistent with our reported NMR data, resulting in significant deviations in the tertiary structure and dynamics from our SARS-CoV-2 s2m model. First, we find differences in the overall three-dimensional structure, where the characteristic 90° L-shaped kink of the SARS-CoV-2 s2m did not form in the Delta s2m resulting in a “linear” hairpin with limited bending dynamics. Delta s2m helical parameters are calculated to align closely with A-form RNA, effectively eliminating a hinge point to form the L-shape kink by correcting an upper stem defect in SARS-CoV-2 induced by a noncanonical and dynamic G:A base pair. Ultimately, the shape difference rationalizes the migration differences in reported electrophoresis experiments. Second, increased fluctuation of the Delta s2m palindromic sequence, within the terminal loop, compared to SARS-CoV-2 s2m results in an estimated increase of entropy of 6.8 kcal/mol at 310 K relative to the SARS-CoV-2 s2m. The entropic difference offers a unique perspective on why the Delta s2m homodimerizes less spontaneously, forming fewer kissing dimers and extended duplexes compared to SARS-CoV-2. In this work, both the L-shape reduction and palindromic entropic penalty provides an explanation of our reported in vitro electrophoresis homodimerization results. Ultimately, the structural, dynamical, and entropic differences between the SARS-CoV-2 s2m and Delta s2m serve to establish a foundation for future studies of the s2m function in the viral lifecycle